1-dehydro-11-keto-progesterone and 17-acyloxy derivatives thereof



United States Patent O l,I-EEHYDRO-ll-KETO-PROGESTERONE AND 17- ACYLOXY DERIVATIVES THEREOF Robert C. Meeks, Kalamazoo, Peter D. Meister, 'Kalamazoo Township, Kalamazoo County, and Herbert C.

Murray, Barry Township, Barry County, Mich., as-

, signors to The Upjohn Company, Kalamazoo, Mich.,

a corporation of Michigan No Drawing. 7 Filed Nov. 2, 1956, Ser. No. 619,943 3 Claims. (31. est-397.45

This; invention relates to a new chemical compound,

Patented Sept. 13, 1960 one are prepared by acylation methods employed in the l-dehydro-l l-ketotestosterone and to the l7 acyloxy de- 1 rivatives thereof, which compounds possess valuable anabolic and androgenic activity.

'The compounds of this invention have the following structural formula:

wherein R is a member of the group consisting of hydrogen and the acyl group of a hydrocarbon carboxylic acid of from one to twelve carbon atoms, inclusive.

In addition to its valuable pharmacological properties mentioned above, the free 17-hydroxy compound of this invention is useful as a starting material for the preparation of l-dehydroadrenosterone, which possesses androgenie and anabolic activity. The free l7-hydroxy group can be oxidized to the 17-keto group through the action of chromic acid. The reaction is conveniently carried out in solution in an organic inert reaction medium, adding at least a molar equivalent of'sodium chromate, preferably an excess, and a mineral acid such as. sulfuric acid. The product, l-dehydroadrenostreone, can be recovered from th'e'reaction medium by evaporation to dryness followed by extraction of 'the residue with methylene chloride followed byrec'rystallization steps to obtain the desired purity.

The novel free 17-hydroxy compound of the invention can be produced by the 'fermentative action of Hypomyces haematococcus var; calicri (Eusar ium solam' var. striatum) .on a1lopregnane-3,11,20 tiione"or by'the fermentative l-dehydrogenation of ll-ketoprogresterone by Hypomyces haematococcus or by a fungus of the genus Septomyxa. The fermentations are each carried out under aerobic conditions; the operational conditions and reaction procedure and details of production can be those already known in the art of steroid bioconversion by US. Patent 2,602,769, utilizing however, respectively, the action of Hypomyces haematococcus var. cancri or a species of the fungus of the genus Septomyxa. Among the species of the former useful in the conversion of allopregnane-3,1l,20-trione is Hypomyces haematocaccus var. cancri, Wollenweber I, Centraalbureau voor Schimmelcultures, Baarn, Holland. Among the species of the art to produce 17-acy1ates from l7-hydroxy compounds of the audrostane series. A suitable method is to dis solve l-dehydro-ll-ketotestosterone in dry pyridine and add an acylating agent to the solution, for example, an anhydride or acid halide of a hydrocarbon carboxylic acid of from one to twelve carbon atoms, inclusive. The 17-acyloxy product is recoverable from the reaction mix? ture by extraction with an organic solvent such as a mixture of ether and chloroform or ether alone. The

' 17-acylates may be recovered in purified form by evaporating the extracting solvent and recrystallizing the residue from acetone-hexane mixture, methanol, or similar recrystallization solvents. Of the 17-acyloxy derivatives included in the compounds of this invention, the 17- acetoxy compound is preferred.

The following examples are illustrative ofthe pro duction of the novel compounds, and are not to be construed as limiting.

EXAMPLE 1 I Y I-dehydro -JJ-ketotest0ster0ne from all0pregnane-3,11,20-

' zrione Twelve liters of a medium containing one percent commercial dextrose hydrate, two percent cornsteep liquor (sixty percent solids), tap water and a small amount of lard oil-'octadecanol anti-foamer was adjusted to pH 4.8 by the addition of sodium hydroxide solution. The medium was sterilized for a period of sixty minutes at twenty pounds steam pressure, at the end of which time the pH was 5.0. The sterilized medium was allowed to cool to 28 degrees centigrade and was inoculated with spores of Hypomyces haematococcus var. cancri (Wollenweber I). Sterile air was admitted through a sparger at the rate of 0.5 liter per minute and the medium agitated with a stirrer at 300 rpm. After a growth period of 48 hours at 28 degrees centigrade, 3.0 grams of allopregnane-3,l1,20-trione dissolved in 130 milliliters of hot ethanol was added. Fermentation was allowed to continue for an additional period of forty hours. The final pII-I was 7.5.

At the end of the fermentation the mycelium was separated from the beer and each was extracted thoroughly with methylene chloride. The combined extract was washed with two percent sodium bicarbonate solution and this was followed by a Water wash. The methylene chloride extract solution was dried with anhydrous sodium sulfate and evaporated at reduced pressure to yield a dry residue weighing 3.4 grams.

An aliquot of the extract taken before evaporation was analyzed by paper chromatography and was found to contain no unconverted allopregnane-3,1l,20-trione.

The residue was dissolved in benzene and separated into its constituents by chromatographic methods. Elution was accomplished with -milliliter portions of solvent as shown in Table I.

ABLE I Fraction Volume Solvents Wt.

360 Benzene 240. 8 240 Benzene-% Ether 100.0 240 Benzene-10% Ethe 48.7 240 Benzene% Ether" 26.0 240 Benzene50% Ether" 25. 7 120 Ether 15. 6 120 Ether '15. 6 120 Etherl0% Ohloroform. 24. 0 120 Ether-10% Ohloroform 74. 6 120 Ether% Chloroform 139.3 120 Ether20% Ohlor0iorm 93. 4 120 Ether-60% Chl0roiorm 37. 4 120 Ether60% chloroform; 7.0 120 Chloroform l. 8 120 Chloroform 274. 8 120 Ohloroform-50% Acetone. 1003.5 120 Ohlor0f0rm50% Acetone-.. 16.0 240 Acetone 20. 5 120 Acetone50% Me OH. 115. 4 120 Methanol 26. 5

:Fraction 21 was dissolved in a mixture of 30:1 methanol-chloroform, filtered, concentrated to two milliliters, and diluted with an equal volume of ether to yield 783 milligrams of crystals having a melting point of 232 to 235 degrees centigrade. Recrystallization from methanol, followed by recrystallization from ethyl acetate, yielded pure l-dehydro-ll ketotestosterone, M.P. 233.5 to 2.34.5 degrees centigrade. It had the following physic properties: [a] +169 degrees 512'; 239 u 7 e 14,600. Infrared spectrum indicates absorption as follows: OH, 3430 cmr nonconjugated ketone, 1695 cm:- h -3-ketone, 1616, 1603 emf in nujol and 1617, 1602 cm." in chloroform; and a conjugated ketone, 1657 cmr i Analysis.Calculated for C H O C, 76.03; H, 7.95. Found: C, 75.97; H, 8.05.

EXAMPLE 2 V l dehydro-ll-ket0t6st0sterone from ll-ketoprogesterone Substantially the same fermentation conditions were followed but employing Septomyxa afiinis, A.T.C.C. 6737 as the organism and ll-ketoprogesterone as the substrate, yielding l-dehydro-ll-ketotestosterone as one of the products separable by chromatographic methods which possessed physical properties agreeing with the product of Example 1, above.

EXAMPLE 3 J-dehydro-Il-ketotestoster0ne 17-acetate A reaction mixture was made by dissolving 83.5 milligrams of l-dehydro-ll-ketotestosterone in 1.0 milliliter of dry pyridine and 0.5 milliliter of acetic anhydride. The solution was kept at about twenty to 25 degrees centigrade overnight, whereupon it was diluted with ice water and extracted with ether. The ether extract was washed with five percent sodium bicarbonate solution; then it was washed with water, and both washing operations were repeated in respective sequence. The ether phase 'of the washed mixture was dried with anhydrous sodium sulfate and concentrated to give 82.5 milligrams of a crystalline residue of crude l-dehydro-l l-ketotestosterone 17-acetate.

The residue was recrystallized from acetone-Skellysolve B hexanes to yield 71 milligrams of crystals having a melting point of 234 to 237 degrees centigrade. Recrys- 5 tallization from acetone yielded pure crystals of l-dehydro-ll-ketotestosterone 17-acetate, MP. 230 to 232 de grees centigrade. It had the following characteristics: [a] -|l35 degrees in chloroform (0.91 cone.). Infrared spectrum analysis confirmed the structure.

Analysis.Calculated for C H O C, 73.66; H, 7.66. Found: C, 73.83; H, 7.64.

Substituting propionic anhydride, benzoyl chloride, decanoyl chloride, octanoyl chloride, valeryl bromide, succinic anhydride and'cyclopentylpropionic anhydride for 15 the acetic anhydride' in. the above procedure 'is productive of the corresponding 17-es'ters of 1-dehydro-1l-ketotestosterone, namely, the 17-propionate, the 17 benzoate, the 17-decanoate, the 17-octanoate, the l7-valerate, the 17- succinate, and the 17-cyclopentylpropionate.

20 l-dehydroel l-ketotestosterone and the 17-acyloxy iderivatives thereof of this invention exhibitvaluable anabolic and androgenic activity and can be administered in the form of tablets, capsules, syrups and the like for oral use or in suitableconventional suspension mediator injection 25 use.

- It is to be understood that the invention is not to be limited to the exact details .of operation or exact compounds shown and described, as obvious modifications and equivalents will be apparent to one skilled in the art, and 30 the invention is therefore to be limited only by the scope of the appended claims.

We claim: 1. A compound of the structural formula:

gen and the acyl group of a hydrocarbon carboxylic acid of from one to twelve carbon atoms, inclusive.

7 2. l-dehydro-ll-ketotestosterone.

3. l-dehydro-ll-ketotestosterone 17-acetate.

References Cited in the file of this patent V V UNITED STATES PATENTS 7 2,756,179 Fried July 24, 1956 2,781,368 Heyl Feb. 12, 1957 2,837,464 Nobile June 3, 1958' 2,902,410 Weintraub et al. Sept. 1, 1959 2,902,498 Fried et a1. Sept. 1, 1959 OTHER REFERENCES H. I. Ringold et al.: J.

I Org. Chem, vol. 21, pages 239- 240, February 1956. l 

1. A COMPOUND OF THE STRUCTURAL FORMULA: 